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首页> 外文会议>Protein Engineering Summit. >Measurement of Fc-Mediated ADCC and CDC Activity of Anti-TNFα and Anti-VEGF Therapeutic Antibodies Using Reporter-Based Bioassays and Engineered TNFα+ and VEGF+ Target Cells
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Measurement of Fc-Mediated ADCC and CDC Activity of Anti-TNFα and Anti-VEGF Therapeutic Antibodies Using Reporter-Based Bioassays and Engineered TNFα+ and VEGF+ Target Cells

机译：使用基于报道的生物测定和工程化TNFα+和VEGF +靶细胞测量抗TNFα和抗VEGF治疗抗体的FC介导的ADCC和CDC活性的测量

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Fc-mediated effector functions are critical to the efficacy and safety of therapeutic antibodies. Measurement of Fc-mediated antibody-mediated cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) during antibody drug discovery and development is not only important for antibodies that harness ADCC and/or CDC as their primary mechanism of action (e.g. rituximab, trastuzumab), but also for antibodies designed to target and block soluble ligands such as TNFa and VEGF. We previously reported the development of a cell-based reporter bioassay platform which has been used to measure ADCC and ADCP mediated through FcγRI, FcγRIIa and FcyRIIIa. These reporter bioassays exhibit the specificity, accuracy, precision, and robustness necessary for qualification according to ICH guidelines and have been used extensively to characterize and measure the potency of antibody-based biologies drugs that target cell surface immune receptors. In the current study, we sought to evaluate Fc-mediated ADCC and CDC activities of therapeutic antibodies designed to target and block soluble ligands including TNFa and VEGF. To measure ADCC activity of anti-TNFa and anti-VEGF blocking antibodies, we developed engineered target cells that express either membrane-bound TNFα or VEGF. When used as target cells with reporter-based effector cells expressing a relevant FcγR, ADCC activity of adalimumab (anti-TNFα) and bevacizumab (anti-VEGF) was detected in a specific and dose-dependent manner. Similarly, when used in a luminescence-based CDC assay, the engineered target cells elicited an appropriate FcγR-mediated response. The assay signals demonstrated IgG isotype specificity as IgG4 variants showed minimal activity in both ADCC and CDC assays. Our results demonstrate that the combined use of cell-based reporter bioassays with target cells engineered to express membrane-bound soluble ligands can provide a simple, specific, and quantitative platform to measure Fc-mediated effector functions of therapeutic antibodies targeting soluble ligands.
机译：FC介导的效应功能对治疗性抗体的功效和安全性至关重要。在抗体药物发现和发育期间的Fc介导的抗体介导的细胞细胞毒性（ADCC）和补体依赖性细胞毒性（CDC）的测量不仅适用于利用ADCC和/或CDC作为其主要作用机制的抗体（例如Rituximab， Trastuzumab），还用于设计用于靶向的抗体，并阻止诸如TNFA和VEGF的可溶性配体。我们之前报道了一种基于细胞的报告生物测定平台，用于测量通过FCγRI，FCγRIIA和FcγRIIA介导的ADCC和ADCP。这些记者生物测定剂根据ICH指南表现出资格所需的特异性，准确性，精度和稳健性，并已广泛使用，以表征和测量靶向细胞表面免疫受体的抗体的生物药物的效力。在目前的研究中，我们寻求评估FC介导的ADCC和CDC活性，所述治疗抗体设计为靶向和阻断包括TNFA和VEGF的可溶性配体。为了测量抗TNFA和抗VEGF阻断抗体的ADCC活性，我们开发了表达膜结合的TNFα或VEGF的工程化靶细胞。当用作表达相关FcγR的基于报道的效应细胞的靶细胞时，以特定和剂量依赖性方式检测到ADALimalab（抗-TNFα）和抗-Vacizumab（抗VEGF）的ADCC活性。类似地，当在基于发光的CDC测定中使用时，工程化靶细胞引发了适当的FcγR介导的响应。测定信号证明了IgG同种型特异性，因为IgG4变体在ADCC和CDC测定中显示出最小的活性。我们的研究结果表明，将基于细胞的报告生物测定与工程化的靶细胞联合使用以表达膜结合的可溶性配体的靶细胞可以提供一种简单，具体的和定量平台，以测量靶向可溶性配体的治疗性抗体的Fc介导的效应功能。

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